Download Maintenance and Regeneration of the Nerve Net in Hydra1 The

AMER. ZOOL., 28:1053-1063 (1988)
Maintenance and Regeneration of the Nerve Net in Hydra1
H. R. BODE, S. HEIMFELD, O. KOIZUMI,
C. L . LlTTLEFIELD, AND M. S. YAROSS
Developmental Biology Center, University of California,
Irvine, California 92717
SYNOPSIS. Due to the tissue dynamics, the entire nervous system of a hydra is in a steady
state of production and loss of neurons. Neurons arise by differentiation from the interstitial cells in the ectoderm. Nerve cell intermediates migrate along the body column,
settle and complete differentiation. The type of neuron formed is a function of axial or
regional location in which the differentiation intermediate settles. Another result of the
tissue dynamics is that all neurons are constantly changing their location. As a consequence,
many neurons switch from one type of nerve cell to another in accord with positional
changes.
spersed among the epithelial cells of both
layers throughout the animal. They are
connected with one another both within a
layer and between the two layers by neuronal processes (Fig. 1), thereby forming a
net of neurons (Hadzi, 1909; Burnett and
Diehl, 1964; Lentz and Barrnett, 1965).
These processes lie near the basal end of
each layer just above the mat of muscle
fibers, which are extensions of the epithelial cells. The neuronal processes run
among the epithelial cells forming synapses
with epithelial cells and nematocytes as well
as with other neurons (Westfall et aL, 1971;
Westfall, 1973).
The organization is simple; only two types
of nerve cells can be distinguished on a
morphological basis: ganglion and sensory
cells. They are very similar as Westfall
(1973) has demonstrated that essentially all
nerve cells in hydra exhibit properties of
sensory, motor, and neurosecretory neurons. Yet, they differ in two respects. One
MORPHOLOGY OF THE NERVE NET
The body plan of a hydra is quite simple, is their location (Fig. 1). The ganglion cell
consisting of a head (hypostome and ten- bodies lie near the base of either cell layer,
tacles), body column, and basal disk (Fig. while the sensory cells are located in a more
3). The structure of the body is a hollow peripheral position (e.g., Diehl and Burcylinder whose wall is comprised of two nett, 1964; Lentz and Barrnett, 1965). Secepithelia, the ectoderm and the endoderm, ond, they differ morphologically in that the
separated by a basement membrane, the cell body of a sensory cell is more elongate
mesoglea (Fig. 1). Nerve cells are inter- (Westfall, 1973). Also, the cilium of a sensory cell extends from the apical surface of
the nerve cell body towards the external
1
From the Symposium on Nervous System Regenerenvironment, and is surrounded at its base
ation in the Invertebrates presented at the Annual Meet- by a stereociliary complex. In comparison,
ing of the American Society of Zoologists, 27-30
the cilium of a ganglion cell projects from
December 1986, at Nashville, Tennessee.
INTRODUCTION
The nervous system of the freshwater
coelenterate, hydra, is one of the most
primitive in the animal kingdom, consisting simply of a loose net of nerve cells. Due
to the continuous expansion and turnover
of the tissue of the animal, it is also highly
unusual in two other respects. The entire
nerve net is in a steady state of production
and loss of neurons, and each neuron is
constantly changing its location in the
animal. As a consequence of these tissue
dynamics, some of the properties of nerve
cell differentiation are also unusual. In the
following, these properties and the dynamics of the nerve net will be described. How
these properties are involved in the maintenance of the nerve net in the context of
the tissue dynamics in the adult, as well as
in the development of the nerve net during
head regeneration, will be discussed.
1053
1054
H. R. BODE ET AL.
FIG. 1. Longitudinal section of a hydra showing the
two tissue layers and the location of the nerve net.
The two enlargements of the regions indicated by the
respective boxes illustrate the location of the nerve
cell bodies and their connecting processes within the
tissue layers. All cell types except nerve cells and epithelial cells have been omitted. The mesoglea is the
basement membrane separating the two layers.
the lateral side and does not have a ciliary
complex (Westfall, 1973).
The nerve net is non-uniform in several
respects. First, 70% of all neurons are in
the ectoderm (Epp and Tardent, 1978). Of
more importance, there is considerable
variation both in the type and density of
neurons found in different regions. Ganglion cells are found throughout the body,
but their density varies considerably
between regions. It is fairly constant
throughout the body column, but increases
rapidly at the apical end reaching on average a 6-fold higher density in the head
(Bode et al., 1973). In the areas where the
tentacles join the hypostome, the density
is still higher forming clusters which
approximate primitive ganglia (Kinnamon
and Westfall, 1981). At the basal end the
density rises gradually down the peduncle
resulting in a 2-4-fold increase in the foot
(Bodega/., 1973; Epp and Tardent, 1978).
There are two types of sensory cells, each
restricted to a specific region of the animal.
The epidermal sensory cells are found only
in the ectoderm of the head. Each sensory
cell body is completely enveloped by an
epithelial cell, and extends from the base
to the apex of that epithelial cell (Fig. 1)
with the cilium projecting into the surrounding environment (Westfall and Kinnamon, 1978). Further, as shown in Figure
2, the epidermal sensory cells exhibit a specific spatial distribution within the head.
In the tentacles they are uniformly distributed with one epidermal sensory cell per
epithelial cell, whereas in the hypostome
there are fewer and all are clustered at the
apex surrounding the mouth (Kinnamon
and Westfall, 1981; Dunne et al., 1985). In
contrast, the other type of sensory cell is
found in the endoderm of the body column, and differs from the epidermal sensory cell in two respects (Davis, 1972). First,
the cell body lies between neighboring epithelial cells instead of being enveloped by
a single epithelial cell. Second, the apical
end of the nerve cell body and cilium extend
towards but do not reach the surface of the
endoderm (Fig. 1).
Recently the use of antibodies has
revealed additional complexity to the nervous system. Grimmelikhuijzen found that
antisera to six different neuropeptides
(FMRFamide, CCK, bombesin, Substance
P, neurotensin, and oxytocin/vasopressin)
each bound to a different subset of nerve
cells (reviewed in Grimmelikhuijzen, 1984).
In each case the subset was located in a
particular part of the head, and in some
cases also in or near the foot. For example,
an antiserum to FRMFamide bound to
nerve cells of the tentacles, hypostome, and
the lower end of the peduncle (Fig. 3). Similarly, monoclonal antibodies have been
generated which bind to different subsets
of the ganglion cells (Bode et al., 1985;
Yaross et al., 1986) as well as to subsets of
the epidermal sensory cells (Koizumi and
Bode, 1988). Thus, the two types of neu-
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HYDRA NERVE NET
FIG. 2. Distribution of ganglion cells (circles) and
epidermal sensory cells (triangles) in the tentacles and
hypostome. One tentacle shown in full, while others
have been truncated.
rons defined by morphology are composed
of subsets that differ in antigenic expression.
BODY X
COLUMN
DYNAMICS OF THE NERVE NET
In adult hydra the epithelial cells of the
body column are continuously undergoing
cell division (Campbell, 1967a; David and
Campbell, 1972). The size of the animal,
however, remains unchanged, as tissue is
continuously lost from the body column by
sloughing at the extremities, or is shunted
into developing buds (Campbell, 19676;
Otto and Campbell, 1977). As illustrated
by the arrows in Figure 3, a result of this
steady state of production and loss is that
individual epithelial cells of both layers are
continually changing their axial location
(Campbell, 19676; Wanek and Campbell,
1982).
Since the nerve net is intertwined among
the epithelial cells, logically one would
expect it to undergo a similar behavior.
Individual nerve cells would be continually
changing their location within the animal
as they are displaced, for example, up the
body column onto a tentacle, along the
tentacle, and eventually lost at the end.
Recently, direct evidence for this was
obtained using a monoclonal antibody, JD1,
which is specific for the epidermal sensory
cells (Dunne et al, 1985). As epithelial cells
are displaced onto the tentacle from the
body column in normal animals, new epidermal sensory cells are added into the net,
usually one with each epithelial cell (Dunne
PEDUNCLE
— BASAL DISK
FIG. 3. Overall structure of a hydra and the distribution of the subset of neurons exhibiting FRMFamide-like immunoreactivity. The regions of the animal
are identified by name. Arrows running along the
body column indicate the direction of tissue displacement. The location of the FRMFamide neurons and
their processes are indicated by the network of dots
and connecting lines.
et al., 1985). This results in the observed
steady-state distribution of JD1 + neurons.
If, however, no nerve cell precursors were
present, no new sensory cells could be
added. With time the distribution of epidermal sensory cells would change as the
nerve net is displaced with the epithelial
cells from tentacle base to tentacle tip. As
illustrated in Figure 4, one would expect a
gap to appear in the pattern of JD1 + sensory cells at the tentacle base shortly after
removal of the precursors. In time the gap
should increase in size, and eventually the
tentacle would be devoid of cells stained
with the antibody.
Yaross et al. (1986) carried out this experiment and exactly this pattern was
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H. R. BODE ET AL.
v-/
Treat with
HU or NM
to remove
nerve cell
precursors
3 days
after treatment
6 days
after treatment
10 days
after treatment
FIG. 4. Experiment illustrating the displacement of epidermal sensory neurons (triangles) along the tentacles
with time in the absence of new nerve cell differentiation.
observed. Further, they observed that the
rate of displacement of epithelial cells and
nerve cells along the tentacle was the same,
providing evidence that the two cell types
move in concert. Since the sensory cells are
an integral part of the nerve net, it is most
likely that all neurons of the net are displaced at the same rate as their surrounding epithelial cells towards one of the
extremities and eventually sloughed.
The important qualitative question in the
face of the continuous displacement of
neurons is how are the regionally specific
subsets of neurons defined by morphology
or antigenic expression maintained? The
explanations of both the quantitative and
qualitative aspects lie in the details of nerve
cell differentiation, which will be described
in the next two sections.
Nerve cell differentiation pathway
GENERATION OF NEURONS AND THE
MAINTENANCE OF THE NERVE NET
The continuous expansion of tissue and
resulting displacement of individual neurons raise important problems as to how
the nerve net is quantitatively maintained
in adult hydra. (1) The loss of neurons by
sloughing at the extremities is continuous,
and yet, the size of the nerve cell population remains unchanged. This suggests that
new neurons must be constantly produced
to maintain their population. (2) In the body
column the density of neurons (number of
neurons/epithelial cell) remains constant
even though new epithelial cells are constantly being added by cell division. Therefore, new neurons must be inserted into
the net throughout the column at a similar
rate to prevent their dilution. (3) The neuron density is higher in the head and foot
than in the body column. Simple displacement of the nerve net from the body column into the head and foot cannot account
for this. The increase in densities must arise
by adding neurons into the net at higher
rates in the extremities than in the body
column.
Nerve cells in hydra, as in other animals,
do not undergo cell division. Pulse-labelling experiments using [3H]thymidine indicate that new nerve cells arise by differentiation throughout the animal (David and
Gierer, 1974; Yaross and Bode, 1978).
Hence, new nerve cells are constantly being
produced. Calculations suggest the rate of
production in sufficient to balance the rate
of loss, and thereby maintain the steady
state (David and Gierer, 1974). The constant tissue growth in hydra as well as the
continuous production of neurons indicates that the nerve cell precursor has the
properties of a stem cell.
Morphological and ultrastructural studies indicate that neurons arise by differentiation from the interstitial cells
(reviewed by Davis, 1974). It is assumed
that neurons are derived from the multipotent stem cells among the population of
large interstitial cells (David and Gierer,
1974; David and Murphy, 1977; Yaross and
Bode, 1978). A large interstitial cell committed to nerve cell differentiation undergoes one or more cell divisions to form
smaller interstitial cells. These are morphologically very similar to neurons, dif-
HYDRA NERVE NET
fering only in the absence of neural processes (Heimfeld and Bode, 1984a, b). They
also resemble intermediates in the nematocyte pathway (David, 1973). The nerve
differentiation intermediates are distinguished from the nematocyte precursors
in that the latter always occur in syncytial
clusters of 8, 16 or 32 cells (Slautterback
and Fawcett, 1959; David and Gierer,
1974), while the former occur as single cells
or in pairs (Bode and Chow, in preparation). As the little interstitial cells of the
nerve cell pathway are also capable of one
or more cell divisions (Bode and Chow, in
preparation), the number of nerve cells
arising from a single committed large interstitial cell is at least 4, and possibly more.
The quantitative distribution of
neuron differentiation is
position-dependent
The most important quantitative feature
of the maintenance of the nerve net that
must be explained is the regional differences in the density of the nerve net. The
initial answer is simple. There is a striking
correlation between the axial distribution
of nerve cells and the spatial pattern of
nerve cell differentiation. The rate of nerve
cell differentiation is high in the head, low
in the body column, and intermediate in
and near the basal disk (Yaross and Bode,
1978). This corresponds directly with the
relative densities of nerve cells in these
regions (Bode et al., 1973). Hence, the
regional pattern of densities is the result
of different levels of neuron differentiation
in the several regions.
This observation leads to the more
intriguing question as to what is the nature
of this position-dependent pattern of differentiation. There are two straightforward explanations. First, this might reflect
a pattern of position-dependent commitment of the stem cells. They would "read"
a positional cue and differentiate accordingly. Or, the pattern could be due to the
selective migration of large interstitial cells
already committed to nerve differentiation, and/or the migration of the differentiation intermediates. Although the
position-dependent commitment of inter-
1057
stitial cells was initially assumed to be the
answer (e.g., Bode and David, 1978), recent
evidence indicates the second explanation
is more probable.
Both large and small interstitial cells are
capable of migration as single cells, and
possibly in pairs (Tardent and Morgenthaler, 1966; Campbell, 1967c; Herlands
and Bode, 1974; Heimfeld and Bode,
1984a). The number that migrate are substantial (Heimfeld and Bode, 1984a). On
average 250 large interstitial cells and 350
small interstitial cells emigrate from any
region of the body column (a region is a
sixth of the column length) within a day.
For the large interstitial cells this amounts
to 15% of their total in any region. Also,
for both large and small interstitial cells
migration is biased since 2-3 times as many
cells move in an apical direction as in a
basal direction. The numbers migrating are
sufficient to account for the observed levels
of nerve cell differentiation.
Some of these migrating cells must be
nerve cell precursors since nerve cells
derived from the migrating cells appear
within 8 hr of emigration (Heimfeld and
Bode, 1984a; Fujisawa, personal communication). Several pieces of evidence indicate that the small interstitial cells are the
precursors. One is the distribution of emigrated cells. The rate of migration of small
interstitial cells is greater than that of the
large ones resulting in a faster and larger
accumulation of the smaller ones in the
head region within 24 hr. Their rate of
accumulation is correlated with the rapid
rise in nerve cells in the head (Heimfeld
and Bode, 1984a). Second, when the cell
composition of the host is altered, the rates
of large interstitial cells immigrating into
the host is unaffected. In contrast, different cell compositions result in corresponding 2-3-fold differences in both the rate of
immigration of small interstitial cells and
subsequent nerve cell differentiation
derived from migrating cells (Heimfeld and
Bode, 19846). Finally, there is a correlation
based on the labelling indices of the
migrating cells and subsequently derived
nerve cells. Pulse-labelled migrating large
and small interstitial cells had labelling
1058
H. R. BODE ET AL.
indices of 53-56% and 61-64%, respectively, while the derived nerve cells had an
index of 63% (Heimfeld and Bode, 19846).
Since single small interstitial cells capable of cell division are known to be intermediates in the nerve cell pathway (Bode
and Chow, in preparation), a fraction of
the migrating small interstitial cells are
most likely such intermediates. Calculations based on the numbers of migrating
small interstitial cells and subsequently
formed nerve cells suggest that the fraction
is greater than 50% of this migrating population. This does not exclude the possibility that some of the migrating large
interstitial cells may also contribute to nerve
cell formation. Indeed, the migrating large
interstitial cell population contains half as
many stem cells as do the non-migrating
large interstitial cells indicating that the
majority of these migrating cells are already
committed to a particular differentiation
pathway (Heimfeld and Bode, 19846).
Thus, the simplest view as to how the
position-dependent pattern of nerve cell
differentiation occurs is the following. Stem
cells among the large interstitial cells
become committed to nerve cell differentiation anywhere along the body column.
A committed cell divides one or more times
to form small interstitial cells which are
nerve cell differentiation intermediates.
These cells migrate and eventually settle
somewhere along the body column or
accumulate in the extremities, where they
finish differentiating into nerve cells.
Accumulation in the extremities accounts
for the higher densities found there as
compared to the body column, and the preferred apical direction of migration
accounts for the highest density being in
the head.
PLASTICITY OF THE DIFFERENTIATED
STATE OF NERVE CELLS
The foregoing described the production
and distribution of nerve cells in terms of
numbers but did not address the question
as to how a migrating small interstitial cell
forms a particular type of nerve cell. In
principle, there are two alternatives. In one,
the differentiation fate (e.g., sensory cell,
or specific neuropeptide expression) of
a migrating small interstitial cell may
have been determined before it began
to migrate. Once determined, it would
migrate selectively to the region where that
particular type of neuron occurs and would
differentiate there. In the other alternative, a migrating small interstitial cell may
be committed to nerve cell differentiation
in general, but not to a particular type of
neuron. Its final location would determine
the type, and would, therefore, be a position-dependent decision. Experiments
based on a consideration of how a specific
subset of neurons arises in the context of
the dynamics of the nerve net indicate the
latter alternative is probably correct.
Plasticity of neuropeptide expression
Two facts lead directly to the idea that
neurons are not irreversibly committed to
a particular differentiated state, which is
the heart of the issue. First, each nerve cell
is continually changing its position in the
animal in concert with the pattern of displacement of the two epithelial layers. Second, the spatial distribution of a subset of
neurons is constant. Thus, at any time one
can demonstrate that the subset of neurons
defined, for example, by their immunoreactivity with an antiserum against
FRMFamide, occur throughout the head,
but are not found in the upper body column (Grimmelikhuijzen et al., 1982;
Koizumi and Bode, 1986). To maintain this
constant pattern, nerve cells exhibiting FLI
(FRMFamide-like immunoreactivity) must
be continuously added at the base of the
tentacles and hypostome to replace the ones
that have been recently displaced further
out on the tentacle, or up the hypostome.
Otherwise the distribution of FLI + neurons would change in time as was observed
(Fig. 4) for the JD1+ epidermal sensory
cells of the tentacles after removal of the
nerve cell precursors (Yaross et al., 1986).
The new FLI+ neurons that are added
to the net could arise by differentiation
from interstitial cells, or by conversion of
FLI" neurons. The evidence suggests the
latter possibility is correct. All nerve cell
precursors (large and small interstitial cells)
can be removed from an animal by treating
it with nitrogen mustard or hydroxyurea
1059
HYDRA NERVE NET
Treat with
HU or NM
to remove
nerve cell
precursors
Decapitate
nerve cell
precursors
Head
regeneration
FIG. 5. Experiment illustrating the conversion of FLI~ neurons to FLI+ neurons during head regeneration.
Circles and triangles represent FLI+ ganglion and FLI+ epidermal sensory cells respectively.
(Diehl and Burnett, 1964;Bode etal., 1976).
When such animals are examined for FLI
binding for up to 3 wk after removal of
the precursors, the spatial distribution of
FLI+ neurons is the same as in normal
animals (Koizumi and Bode, 1986). Were
maintenance of the pattern dependent on
new differentiation, changes in the distribution with time would have been expected
as in Figure 4. The constancy of the pattern suggests that FLI~ neurons are converted into FLI+ neurons as they are displaced from the upper body column onto
a tentacle or the hypostome.
Another experiment, illustrated in Figure 5, provided more direct evidence that
changing the position of a neuron could
change its neuropeptide expression.
Removal of the head results in the regeneration of a new head by the tissue at the
apical tip of the decapitated animal. The
process is morphallactic in that it involves
the remodeling of the existing tissue and
is independent of cell division (Hicklin and
Wolpert, 1973; Cummings and Bode,
1984). Thus, epithelial cells and neurons
formerly in the body column become part
of a tentacle or the hypostome, and have
in effect changed their axial location. Animals devoid of all nerve cell precursors were
bisected in the upper body column to
remove the head and all the FLI+ neurons
of the apical end of the animal. Following
regeneration of a head they once again
exhibited FLI+ neurons (Koizumi and
Bode, 1986). Such neurons could only have
arisen from either FLI~ neurons or epithelial cells since these were the only two
cell types in the ectoderm. As epithelial
animals, which consist only of epithelial
cells, do not form nerve cells (Marcum and
Campbell, 1978) the reappearance of the
FLI + neurons must be due to the conversion of FLI" neurons of the body column
into FLI + neurons as they are incorporated into the head after tissue reorganization. Hence, a change in the location of
a neuron can alter its neuropeptide expression. The same has also been demonstrated
for the subset of neurons expressing vasopressin-like immunoreactivity (Koizumi and
Bode, in preparation).
These results raised a second question:
Is the reverse possible? Can FLI + neurons
turn off FLI expression if they enter a FLI~
region? Normally neurons are displaced
from the FLI" body column basally into
the FLI+ lower peduncle, and later into the
basal disk, which is again FLI" suggesting this reversal might occur. This possibility was examined by transplanting
lower peduncle regions into the middle of
the body column. In all grafts in which the
transplanted tissue took on the character
of the body column, FLI disappeared. In
those grafts in which the transplanted tissue maintained its lower peduncle character, FLI+ neurons were observed
(Koizumi and Bode, 1986). Apparently
expression of the neuropeptide can be
turned on or off depending on the axial
location of the neuron.
Conversion of ganglion cells
into sensory cells
The reappearance of FLI + neurons in
the regenerated hypostome also suggests
another type of switch in neuron phenotype was taking place. Both FLI + ganglion
and FLI + epidermal sensory cells occur in
the hypostome (Koizumi and Bode, 1986).
As there are no epidermal sensory cells in
1060
H. R. BODE ET AL.
Nv 1
Li
t
Nv 2
I m — Bi
Nv 3
\
Li
Nv4
FIG. 6. Nerve cell differentiation pathway with the
minimum in complexity. Im: multipotent stem cell;
Bi: large interstitial cell committed to nerve cell differentiation; Li: small interstitial cell = differentiation
intermediate; Nvl-Nv4: specific types of neurons.
the body column (Westfall, personal communication), and neurons such as endodermal sensory cells do not move across
the mesoglea (Smid and Tardent, 1984),
the FLI + epidermal sensory cells probably
arose from ganglion cells in the body column ectoderm. The use of two monoclonal
antibodies provided more direct evidence
for this deduction.
The monoclonal antibody, TS33, binds
only to epidermal sensory cells of the
hypostome, and to no other neurons in the
animal. To demonstrate the conversion of
ganglion cells to sensory cells, the same
experimental design described for conversion of FLI~ neurons to FLI + neurons
(Fig. 5) was used. Animals devoid of interstitial cells were decapitated, allowed to
regenerate, and found to exhibit TS33 +
epidermal sensory cells in the regenerated
hypostome (Koizumi and Bode, 1988).
Because of the absence of sensory cells in
the body column ectoderm, TS33 + epidermal sensory cells must have arisen from
TS33— ganglion cells.
Evidence for intermediates undergoing
this transition was obtained by combining
the use of TS33 with a second antibody,
TS26, which binds to ganglion cells in the
body column and the head. If ganglion cells
were being converted to sensory cells during regeneration, one might expect to see
cells in transition that bind both antibodies. The experiment was repeated and
regenerates were double-labelled with the
two antibodies using indirect immunofluorescence and two fluorochromes. On
average, 2 double-labelled epidermal sensory cells were found per regenerate
(Koizumi and Bode, 1988). As regeneration
proceeded this number remained constant,
while the number of TS33+ epidermal
sensory cells per animal increased. This is
consistent with the idea that the doublelabelled cells are in transition from a ganglion to an epidermal sensory cell. After
obtaining this result, careful examination
of normal animals revealed that a small
number of double-labelled sensory cells
were present suggesting that this conversion also occurs normally.
It is plausible that the conversion from
ganglion cells to sensory cells is a normal
maturation process, and not due to a change
in position. The following experiment renders this unlikely. Regeneration of the head
at the apical tip of the lower half of an
animal following bisection in the middle of
the body column resulted in the formation
of epidermal sensory neurons in the new
head (Koizumi and Bode, 1986). As all the
neurons that formed the head were originally destined to be displaced in a basal
direction, none of them would ever have
formed an epidermal sensory cell. Therefore, the appearance of new epidermal sensory cells could not have been due to maturation, but was due to a change in position.
Hence, a consequence of neurons continuously changing their location as the
epithelia are displaced is that they can alter
their differentiated state from one type of
neuron to another. A direct implication of
this is that the type of differentiation
undertaken by a migrating small interstitial cell depends on the region in which it
settles. If it stops in the head, it will "read"
its location, and form, for example, a neuron that expresses FLI + .
CONCLUSIONS
There are now sufficient data to draw a
first approximation of the development and
maintenance of the nerve net. In a normal
adult the net is maintained in a steady state
by the continuous loss of neurons at the
extremities balanced by a constant pro-
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HYDRA NERVE NET
duction of new nerve cells throughout most
of the animal. During a process such as
head regeneration, the net is in part developed anew. Both situations require the
generation and appropriate distribution of
specific numbers and types of neurons.
The current picture as to how neurons
are generated is shown in Figure 6, and
represents the minimum in complexity.
Neurons arise by differentiation from multipotent stem cells in a two-step commitment process. At all times some multipotent stem cells (Im) among the large
interstitial cells undergo a commitment to
form nerve cells, but the type of neuron is
not specified. For two reasons, this initial
commitment is probably due to an internal
mechanism. It can take place anywhere
interstitial cells occur in the animal suggesting that commitment to nerve cells does
not require a particular environment. Also,
there is no evidence for external control
of this step. One can imagine a number of
internal mechanisms ranging from genetically fixed programs to stochastic decisions based on a set of probabilities for
differentiation.
The committed cell (Bi) divides to produce two small interstitial cells (Li), which
are differentiation intermediates that are
also not committed to type of neuron.
These intermediates migrate, and will
eventually settle in the body column or in
one of the extremities. The type of neuron
formed (Nvl -Nv4) is influenced by the final
location of the small interstitial cell. This
is the second step in the commitment process.
The specific distributions of neurons can
be explained in part with the same process.
The asymmetric densities of neurons are a
result of the migration behavior of the differentiation intermediates. Many small
interstitial cells accumulate in the extremities resulting in higher rates of neuron
differentiation there than in the body column. Since migration occurs preferentially
in an apical direction, the highest differentiation rate and density are found in the
head.
The position-dependent distribution of
subsets of types of neurons is due to the
intermediates "reading" their final loca-
tion, and differentiating accordingly. The
constancy of this pattern in the face of the
continuous change of location of each neuron can be attributed to the metastability
of the differentiated state of many of the
neurons. They change their type of differentiation (Nv2 -• Nvl) as they change position during tissue movements. Whether all
neurons are plastic in this regard is not
known. Although many can arise by conversion from other neurons, some arise only
by differentiation from interstitial cells
(Nv3, Nv4). An example is the subset of
epidermal sensory cells in the tentacles,
defined by the monoclonal antibody, JD1,
which was not replaced when the precursors were removed (Yaross et al., 1986).
Finally, the metastability of the differentiated state of many of the neurons in
the hydra nervous system was observed
simply because neurons change position
within the animal. This raises the possibility that the differentiated state of neurons
in other animals may also be metastable
rather than inherently terminal. The differentiated state may only appear to be terminal because neurons in other animals seldom change their location.
ACKNOWLEDGMENTS
We thank Patricia Bode for a critical
reading of the manuscript. Some of the
research described was supported by
research grants from the National Institutes of Health to H.R.B. (HD 08086 and
HD 16440), from the National Science
Foundation to M.S.Y. (PCM-83-02581),
and from the Japanese Ministry of Education and Fukuoka Prefecture to O.K.
Current addresses: Shelly Heimfeld,
Dept. of Pathology, Stanford University,
Stanford, California 94305.
Osamu Koizumi, Physiological Laboratory, Department of Science, Fukuoka
Women's University, Higashi-ku, Fukuoka, 813 Japan.
Marcia Yaross, Pharmacia Intraocular
Intermedics, Pasadena, California 91140.
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